Generation and Use of Recombinant Galectins.

Autor: Wu SC; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts., Paul A; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts., Ho A; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts., Patel KR; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts., Allen JWL; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts., Verkerke H; Center for Transfusion Medicine and Cellular Therapies, Emory University School of Medicine, Atlanta, Georgia., Arthur CM; Center for Transfusion Medicine and Cellular Therapies, Emory University School of Medicine, Atlanta, Georgia., Stowell SR; Joint Program in Transfusion Medicine, Department of Pathology, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts.
Jazyk: angličtina
Zdroj: Current protocols [Curr Protoc] 2021 Mar; Vol. 1 (3), pp. e63.
DOI: 10.1002/cpz1.63
Abstrakt: Galectins are soluble carbohydrate binding proteins that can bind β-galactose-containing glycoconjugates by means of a conserved carbohydrate recognition domain (CRD). In mammalian systems, galectins have been shown to mediate very important roles in innate and adaptive immunity as well as facilitating host-pathogen relationships. Many of these studies have relied on purified recombinant galectins to uncover key features of galectin biology. A major limitation to this approach is that certain recombinant galectins purified using standard protocols are easily susceptible to loss of glycan-binding activity. As a result, biochemical studies that employ recombinant galectins can be misleading if the overall activity of a galectin remains unknown in a given assay condition. This article examines fundamental considerations when purifying galectins by lactosyl-sepharose and nickel-NTA affinity chromatography using human galectin-4N and -7 as examples, respectively. As other approaches are also commonly applied to galectin purification, we also discuss alternative strategies to galectin purification, using human galectin-1 and -9 as examples. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Purification of galectins using lactosyl-sepharose affinity chromatography Basic Protocol 2: Purification of human galectin-7 using a nickel-NTA affinity chromatography column Alternate Protocol 1: Iodoacetamide alkylation of free sulfhydryls on galectin-1 Alternate Protocol 2: Purification of human galectin-9 using lactosyl-sepharose column chromatography.
(© 2021 Wiley Periodicals LLC.)
Databáze: MEDLINE
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